A major objective in this research is to determine how a regulatory gene located on one chromosome controls the expression of genes on other chromosomes. The regulatory system chosen for study is the galactose utilization pathway in yeast. The system consists of four structural genes specifying four inducible enzymes and three genetically well defined regulatory genes which control enzymes inducibility. The immediate aim is to determine how regulatory gene control of inducible enzyme activity is mediated. To accomplish this aim, we propose to use immunoprecipitation and cell-free translation to detect the in vivo and in vitro mRNA-directed synthesis of two of the four inducible enzymes encoded by the structural genes in the system. Well defined mutations mapping at the regulatory gene loci will be used to determie at which level, transcripton, translation or enzyme activation, regulatory gene control is exerted. Since the three structural genes (GAL1, GAL7, GAL10) under study here are tightly linked on chromosome II, and since our preliminary data indicate that induction of enzyme activity is due to de novo synthesis of messenger RNA for at least one of the enzymes, we will also determine whether a polycistronic mRNA is involved in the induction of all three enzymes. The mRNAs specified by the GAL1 and GAL7 genes will be used to screen a recombinant DNA plasmid bank for complementary DNA sequences. The transcriptional organization and regulation of the GAL cluster will be determined.